Closed photobioreactor and method of use

ABSTRACT

An apparatus is disclosed for the controlled production of microorganisms by photosynthesis in a closed photobioreactor. The closed photobioreactor contain a photosynthetic culture in a substantially sealed environment and provides a system for recirculating the reactant gas through the culture. This closed loop system can be operated with expensive carbon isotopes (i.e.,  13  CO 2  or  14  CO 2 ). Also, a system is provided for removing the molecular oxygen produced in the photosynthesis reaction from the closed photobioreactor. Furthermore, a pH-regulated control valve is utilized for controlling the addition of reactant gas to the culture in response to the alkalization of the culture.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The present invention is directed to the controlled production ofmicroorganisms by photosynthesis in a closed photobioreactor containinga photosynthetic culture in a substantially sealed environment andwherein a reactant gas is recirculated through the algal culture.

2. Description of the Prior Art

Algae have been cultivated artificially for such diverse purposes as theproduction of food for animals and humans, the treatment of sewage andwaste waters, and the accumulation of radioactive wastes. More recently,algal cultures have been used for the production of enzymes havingindustrial and research applications and for producing oils and othermaterials having nutritional value. Modern biotechnology offers anopportunity for the genetic modification of algae to yield culturescapable of producing a wide variety of useful materials.

Various methods and equipment have been employed for the artificialculturing of algae. Perhaps the simplest procedures have involved theuse of shallow open ponds exposed to sunlight. Such ponds are subject tocontamination by dust, other microorganisms, insects and environmentalpollutants and provide minimal ability to control the degree of exposureto light, temperature, respiration and other important factors. A moresophisticated approach has involved growing algal cultures inplastic-covered trenches and ponds, optionally having electricallypowered pumps and agitators. These configurations reduce the chances ofcontamination of the culture and permit more accurate control oftemperature, respiration and other parameters.

Modern photobioreactor structures are constructed to optimize thephotosynthetic process by providing a means for uniformly exposing thecells in the algal culture to the optimum amount of visible light. Toaccomplish this, prior photobioreactors have been built with sources oflight, e.g., fluorescent tubes, optical rods etc., mounted in thephotobioreactor, immersed in the algal culture. The light sources arepositioned inside the photobioreactor taking into consideration suchcharacteristics as the cell density and light path length.

The principal nutrient required for the algal culture in thephotosynthesis process is inorganic carbon. In known photobioreactorsystems, the algal cultures obtain their carbon from carbon dioxide,often bubbled through the culture medium. The carbon dioxide is oftenintroduced in the medium through sparging tubes or other suitable meanspositioned near the bottom of the photobioreactors. The bubbling of thecarbon dioxide often serves a dual function in that it aids in thecirculation of the algal culture.

The presently known photobioreactors operate in what could be called anopen-loop mode, that is, there is a free exchange of gases between theatmosphere and the interior of the photobioreactor. Thesephotobioreactors are characterized in that they have open tops or topswhich are not in sealed relation with the tank containing the algalculture. As the photosynthesis process occurs, the gases produced,oxygen being the main by-product of the biochemical transformation, areallowed to escape from the photobioreactor into the atmosphere.

Operating the photobioreactor in this open-loop mode is oftensatisfactory because the materials lost to the atmosphere, i.e., carbondioxide, evaporated water comprising the liquid culture medium, arerelatively inexpensive and are not environmentally harmful. In addition,oxygen produced in the photosynthesis reaction which if contained couldresult in overpressurization, is allowed to freely escape. However, whenvery expensive reactant gases such as carbon isotopes ¹³ CO₂ or ¹⁴ CO₂are used in such systems, economically unacceptable losses result ifthese rare isotopes are allowed to freely escape. Furthermore, ifdeuterium oxide is utilized in the liquid culture medium, excessiveevaporative losses of this expensive material may occur as well.

SUMMARY OF THE INVENTION

The present invention provides a novel photobioreactor system whichovercomes the aforementioned problems and which provides efficient andeconomical operation while enabling the use of expensive reactant gasesand other reactant materials in the system. In one embodiment, the novelphotobioreactor system is operated in a closed loop mode wherein thereactant gas is introduced into the liquid culture medium forphotosynthetically reacting with the photosynthetic culture and isrecirculated through the culture in a substantially sealed environment.The closed photobioreactor system further comprises means for removingthe molecular oxygen produced by the photosynthetic reaction withoutsubstantial leakage of the reactant gas.

The present invention further provides for the use of pH sensing tocontrol the concentration of the reactant gas in the algal culture inresponse to the alkalization of the culture medium. This is accomplishedby monitoring the pH of the culture and actuating control means to admitthe reactant gas to the culture when the pH of the culture exceeds apreselected reference value.

The invention thus enables much more economically efficient operation ofa photobioreactor, as the reactant gas and the other materials are usedin the liquid culture medium are substantially contained within thesystem and thus not lost to the atmosphere. Furthermore, controlling theconcentration of the reactant gas in response to the pH of the algalculture results in efficient use of the reactant gas and preventsoverpressurization when used in the closed photobioreactor system.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a schematic view of a closed photobioreactor system embodyingthe present invention;

FIG. 2 is a schematic view of a closed photobioreactor system inaccordance with a further embodiment of the present invention;

FIG. 3 is a perspective view of an open photobioreactor in accordancewith a further embodiment of the present invention;

FIG. 4 illustrates a device for removing photosynthetically generatedoxygen which enables the oxygen to react with metal filings;

FIG. 5 illustrates the removal of photosynthetically generated oxygen bychemisorption; and

FIG. 6 shows a further embodiment of the present invention which removesphotosynthetically generated oxygen from a reactant gas by an oxygenscrubber.

DESCRIPTION OF THE PREFERRED EMBODIMENTS

Referring now to the drawings, FIG. 1 shows a schematic of a closedphotobioreactor system 10 which includes means for removing themolecular oxygen produced in the photosynthetic reaction and means forcontrolling addition of the reactant gas to the photobioreactor.

As illustrated in FIG. 1, the closed photobioreactor system 10 comprisesa tank 11 for containing a liquid photosynthetic culture 14 in asubstantially sealed environment. The tank,s top portion 18 isconstructed to be sealed with tank 11 so that gases produced in thephotosynthesis reaction, e.g., molecular oxygen and any evaporation fromthe liquid culture, are substantially sealed within the system andprevented from being lost to the atmosphere. Light tubes 12, which canbe for example fluorescent tubes, are positioned in the algal culture toprovide light for the photosynthetic reaction. The light tubes can bepositioned within or above the algal culture or in any way known tothose skilled in the art to effect the photosynthetic reaction. Forexample, the light tubes 12 can be arranged in the photobioreactor asdisclosed in pending U.S. patent application Ser. No. 07/163,800,assigned to assignee of the present application and incorporated hereinby reference.

The reactant gas is introduced to the algal culture through spargingtubes 13, positioned near the bottom of tank 11. Bubbling the reactantgas through the liquid medium also serves to agitate and circulate thealgal culture as illustrated by the arrows in ¹³ CO₂, FIG. 1. Thereactant gas, such as the isotope CO₂, is stored in a tank 21, and isthe reactant gas used in one preferred embodiment of the presentinvention as shown in FIG. 1. Photosynthetically producingmicroorganisms using this rare carbon isotope create a universallylabeled biomass which has many important uses such as in non-invasivediagnostics. For example, magnetic resonance spectroscopy can detect ¹³C in sugars stored in the body and can detect organic C compoundsassociated with various bodily chemical functions. Other suitablereactant gases such as ¹⁴ CO₂ or CO₂ may be utilized to fulfill theculture,s carbon requirement.

The photobioreactor system of FIG. 1 operates in a substantially sealedenvironment and includes means for removing the molecular oxygenproduced by the photosynthesis reaction. Oxygen removal is accomplishedin the embodiment shown by means of a catalytic converter 15, into whichhydrogen is introduced in a controlled manner to react with themolecular oxygen in the presence of a catalyst. The water vapor formedby the combustion reaction is condensed by a condenser 26 and occupies aminimum volume in the form of water. The catalytic converter utilized inthe present invention can be a basic catalytic converter in its "off theshelf" condition. Water formed in the combustion reaction may becollected and removed from the system in any known and conventional waysuch as through a drain 26a.

As illustrated in FIG. 1, the hydrogen introduction into catalyticconverter 15 is controlled by control valve 27 which is activated by arelay 20. Control valve 27 can be solenoid valve or any valve known tothose skilled in the art which can be controlled to pass a desiredamount of gas. Relay 20 receives a signal responsive to oxygenconcentration level from an oxygen sensor 30 and an oxygen responsiveelectrode 23. If oxygen electrode 23 is such that it responds to gasphase oxygen it should be positioned in the region above photosyntheticculture whereby it measures the oxygen concentration level in the gas.If oxygen electrode 23 is such that it responds to dissolved oxygen, theelectrode may be submerged anywhere in the culture. Oxygen electrode 23is electrically connected to oxygen sensor 30 which generates a signalrepresenting the measured oxygen level. The relay 20 then opens thecontrol valve 27 when a preset oxygen level (gas phase or liquid phase)measured by the oxygen electrode 23 is exceeded and hydrogen stored intank 22 is caused to flow into the catalytic converter 15.

To avoid the possible introduction of an excessive amount of hydrogeninto the catalytic converter, sensor and alarm means 15b can be providedto measure the hydrogen concentration flowing from the catalyticconverter and to signal the presence of abnormal amounts. Ideally,hydrogen will be added at a rate sufficient to react with all of themolecular oxygen generated by photosynthesis. Excessive amounts ofhydrogen could result in an explosion or the introduction of thehydrogen into the algal culture. Therefore, hydrogen addition ispreferably controlled such that the combination is stoichiometricallybalanced slightly on the side of excess oxygen.

As discussed above, the hydrogen is reacted with the elemental oxygen ina controlled manner and the photosynthetically generated oxygen isthereby effectively removed from the gases circulating in the tank 14.As further illustrated in FIG. 1, the gases remaining after the oxygenremoval, which will comprise primarily the reactant gas, are pumped by apump 28 back through the algal culture.

Another feature of the present invention involves the controlledaddition of the reactant gas, e.g. CO₂, ¹³ CO₂ or ¹⁴ CO₂, to the algalculture in the closed photobioreactor. As stated above, algal culturestypically obtain their carbon from a gas, e.g. CO₂, which is bubbledthrough the algal culture medium, establishing the following equilibria:

    CO.sub.2 +H.sub.2 O⃡H.sub.2 CO.sub.3 ⃡H.sup.+ +HCO.sub.3 ⃡H.sup.+ +CO.sub.3.sup.-2

A photosynthetic algal culture will consume COz from the mediumresulting in the increased alkalization of the solution. The presentinvention uses pH responsive means, shown in FIG. 1 in the form of apH-regulated control valve 29, to admit the reactant gas to the algalculture responsive to the alkalization level of the culture medium.Control valve 29 is controlled by relay 19 which receives a signalindicating the pH of the algal culture from pH sensor 17 via pHelectrode 24. When the measured pH value exceeds a preselected referencevalue, relay 19 opens the control valve 29 which admits the reactant gasto the culture from the container 21. When the pH of the medium fallsbelow the preselected reference value, the valve is closed and the inputof the reactant gas is cut off.

This aspect of the present invention has several useful applications forthe culturing of photosynthetic algae; for example: (1) the system canregulate the pH of an algal culture; (2) the system can be an integralcomponent of a closed photobioreactor in which the gas stream isrecycled through the culture. Furthermore, it is important that theinput of reactant gas be controlled so as not to overpressurize thephotobioreactor. Adding the reactant gas only in response to the pHchanges of the culture greatly alleviates the danger ofoverpressurization. Additionally, this system is especially useful whena closed photobioreactor is operated with expensive carbon isotopes(i.e. ¹³ CO₂ or ¹⁴ CO₂) since the isotope is only admitted to theculture in response to its consumption by the algae.

In an alternative embodiment of the present invention as shown in FIG.2, molecular oxygen removal can be accomplished by a reaction means 50for chemically reacting the molecular oxygen to form a stable chemicalcompound thereof with at least one additional element. In the particularembodiment shown, the reaction means 50 comprises a fuel cell 50a whichcauses hydrogen from a chamber 51 to react with the molecular oxygen ina controlled manner through an ion exchange membrane 52. The ionexchange membrane can be a hydrated Al₂ O₃ membrane or any suitablemembrane which facilitates ion exchange and the reaction of the hydrogenwith the molecular oxygen. The fuel cell 50a illustrated is of thehydrogen-oxygen type and typically includes a catalyst, such asplatinum, to assist in the reaction. Such fuel cells are well known inthe art. The small amount of electrical energy produced in this reactionmay be carried through wires 53 and converted to heat in resistor 54.The elements in FIG. 2 having the same reference numerals as those inFIG. 1 are the same and perform the same functions as already describedin connection with FIG. 1.

As illustrated in FIG. 2, hydrogen stored in tank 22 is admitted tochamber 51 in the same controlled manner as set forth with reference tothe closed photobioreactor in FIG. 1. Control valve 27 is actuated byrelay 20 when the preset oxygen level measured by oxygen electrode 23 isreached. Also, when the oxygen level falls below the preset level, thehydrogen flow from tank 22 is shut off. The preset oxygen level shouldbe at about 20%, or at about atmospheric concentration, or other desiredconcentration could be maintained. The oxygen removal means 50 also hasmeans for sensing the hydrogen concentration in chamber 51 and triggersan alarm when concentration exceed preset limits. Furthermore, anymembrane system which preferentially allows the diffusion of the O₂ overCO₂ could also be used.

In a further embodiment of the closed photobioreactor of the presentinvention, FIG. 4 illustrates the removal of photosyntheticallygenerated oxygen from a reactant gas by enabling the oxygen to reactwith a metal forming a metal oxide in removal means 40. The reactant gasand the photosynthetically generated oxygen are introduced into removalmeans 40 containing a bed metal filings 41 (or a metal "wool") whichwill react with substantially all the photosynthetically generatedoxygen forming a metal oxide. Oxygen is thus removed from the gasstream. Copper filings heated to approximately 500° C. can be used orany metal known to those skilled in the art which will readily form ametal oxide in the presence of oxygen.

In yet a further embodiment of the closed photobioreactor of the presentinvention, FIG. 5 illustrates the removal to photosyntheticallygenerated oxygen by chemisorption. In chemisorption, which is ashortening of chemical absorption, the molecules stick to the surface ofa metal as the result of a chemical, and usually a convalent, bond. Asshown in FIG. 5, the reactant gas and the photosynthetically generatedoxygen are introduced into the apparatus 55 wherein a solid active metalremoves the oxygen by chemisorption. A device of this type is anOxisorb, manufactured by Analalabs, Inc.

In still a further embodiment of the present invention, FIG. 6illustrates removing photosynthetically generated oxygen from a reactantgas by an oxygen scrubber. Oxygen scrubber 36 utilizes a "BTS catalyst",which is first reduced with H₂ or CO, and placed in the gas stream toeffectuate oxygen removal. Also, "Ridox" which is an active granularreagant can be used in oxygen scrubber 36 to remove the oxygen.

The closed photobioreactor shown in FIG. 6 operates in a mannersubstantially similar to the photobioreactor in FIG. 1. Relay 20receives a signal responsive to the oxygen concentration level fromoxygen sensor 30 and an oxygen responsive electrode 23. Relay 20 opens acontrol valve 44 when a preset oxygen level is exceeded thus introducingthe reactant gas and photosynthetically generated oxygen into oxygenscrubber 36. When the oxygen concentration level is below the presetvalue, the reactant gas flows through control valve 44 and along bypasslines 46 to bypass oxygen scrubber 36. In both cases, the reactant gasis recirculated through the photosynthetic culture as discussed withreference to FIG. 1. Furthermore, additional reactant gas (CO₂, ¹³ CO₂or ¹⁴ CO₂) can be supplied to the culture as shown in FIG. 1.

In addition to the solid oxygen scrubbers described above, a liquidscrubber can be used in scrubber 36 as well to remove thephotosynthetically generated oxygen from the reactant gas. In thisembodiment, the gas stream is bubbled through a liquid containingchemicals which react with oxygen, and thus effectively removing it fromthe gas stream. In a preferred embodiment, this liquid comprises 0.4M Cr(C10₄) in HCl with amalgamated Zn, but any such liquid known to thoseskilled in the art could be used as well.

In a further embodiment of the present invention, FIG. 3 illustrates thepH controlled addition of reactant gas such as carbon dioxide into thealgal culture in an open photobioreactor 60. The control valve 29controls the introduction of CO₂ to the algal culture in response to theculture's alkalization in substantially the same way as described withreference to FIG. 1. Here, however, the reactor container is formed of atank 61 which is open to the atmosphere and the inexpensive gases areallowed to freely escape. The pH-controlled addition of reactant gas tothe algal culture is advantageous because the system can regulate the pHof the culture, and the reactant gas is efficiently used when admittedon as required to satisfy the reaction rate within the culture inresponse to its consumption by algae.

While it is apparent that the preferred embodiment shown and describedprovides certain advantages, it should be understood that suchembodiments are presented for the purpose of making a full, clear anddetailed disclosure thereof and that many of the advantages of thepresent invention can nevertheless be realized in other configurations,and it will be appreciated that various modifications, changes andadaptions can be made, all of which are intended to be comprehendedwithin the scope of the appended claims.

We claim:
 1. A closed photobioreactor comprising:(a) container means forcontaining a photosynthetic algal culture in a substantially sealedenvironment; (b) means for introducing light into said container means;(c) means for introducing a reactant gas into said container means forphotosynthetically reacting with a culture in said container means inthe presence of light such that molecular oxygen is produced as aby-product, the molecular oxygen and unconsumed reactant gas forming agas mixture; (d) means for removing the molecular oxygen from the gasmixture without leaving by-products of the oxygen removal to berecirculated and without substantial leakage of the unconsumed reactantgas; (e) means for recirculating the unconsumed reactant gas afteroxygen removal through an algal culture in the substantially sealedenvironment when the reactant gas and the algal culture are present insaid container means; and wherein said means for removing molecularoxygen comprises means for enabling a combustion reaction with theoxygen and at least one other element to form a stable chemical compoundthereof with said at least one other element.
 2. The closedphotobioreactor as set forth in claim 1 further comprising means forsensing the concentration level of oxygen in said container means. 3.The closed photobioreactor as set forth in claim 2 wherein said meansfor removing molecular oxygen comprises a first chamber for containingthe molecular oxygen, a second chamber for containing hydrogen and anion exchange membrane separating said first and second chambers forenabling the hydrogen when present in said second chamber to react withthe oxygen.
 4. The closed photobioreactor as set forth in claim 3wherein the ion exchange member is made of Al₂ O₃.
 5. The closedphotobioreactor as set forth in claim 3 further comprising means forintroducing the hydrogen into said second chamber.
 6. The closedphotobioreactor as set forth in claim 1 wherein said means for enablinga combustion reaction comprises a catalytic converter.
 7. The closedphotobioreactor as set forth in claim 6 further comprising means forintroducing hydrogen into said catalytic converter.
 8. The closedphotobioreactor as set forth in either claim 5 or claim 7 wherien saidmeans for introducing the hydrogen comprises valve means actuatedresponsive to said means for sensing the concentration level of oxygen.9. The closed photobioreactor as set forth in claim 1 further comprisingmeans for controlling the addition of said reactant gas into saidculture in response to the pH of said culture.
 10. The closedphotobioreactor as set forth in claim 9 wherein said means forcontrolling the addition of said reactant gas comprises pH-controlledvalve means for admitting said reactant gas responsive to the pH of thealgal culture exceeding a predetermined level.
 11. The closedphotobioreactor as set forth in claim 1 wherein said means forintroducing reactant gas is a means for introducing carbon dioxide. 12.The closed photobioreactor as set forth in claim 1 wherein means forintroducing a reactang gas is a means for introducing ¹³ Co₂ or ¹⁴ CO₂.13. A method of performing a continuous photosynthetic reaction processcomprising:(a) maintaining a photosynthetic algal culture in asubstantially sealed environment; (b) circulating a reactant gas throughsaid culture in the presence of light and photosynthetically reactingsaid gas with said culture and thereby producing molecular oxygen insaid reactant gas as a by-product, the molecular oxygen and unconsumedreactant gas forming a gas mixture; (c) removing the molecular oxygenproduced after photosynthetically reacting the reactant gas with saidculture, from the gas mixture, said removing the molecular oxygencomprising chemically reacting said oxygen with at least one additionalelement to form a stable chemical compound therewith and without thestable chemical compound or other by-product the chemical reaction beingrecirculated into said algal culture and wherein said removing saidmolecular oxygen comprises introducing said molecular oxygen into afirst chamber, introducing hydrogen into a second chamber and reactingsaid molecular oxygen with said hydrogen through an ion exchangemembrane separating said first and second chambers to form water as atleast one by-product; and (d) recirculating the unconsumed reactant gasafter oxygen removal into said algal culture in said substantiallysealed environment.
 14. The method of performing a continuousphotosynthetic reaction process as set forth in claim 13, furthercomprising the step of controlling the introduction of hydrogen intosaid second chamber.
 15. The method of performing a continuousphotosynthetic reaction process as set forth in claim 14 furthercomprising sensing the level of oxygen concentration in said reactantgas.
 16. The method of performing a continuous photosynthetic reactionprocess as set forth in claim 13 wherein introducing a reactant gas intosaid algal culture comprises introducing Co₂ or ¹³ CO₂ or ¹⁴ CO₂.
 17. Amethod of performing a continuous photosynthetic reaction processcomprising:(a) maintaining a photosynthetic algal culture in asubstantially sealed environment; (b) circulating a reactant gas throughsaid culture in the presence of light and photosynthetically reactingsaid gas with said culture and thereby producing molecular oxygen insaid reactant gas as a by-product, the molecular oxygen and unconsumedreactant gas forming a gas mixture; (c) removing the molecular oxygenproduced after photosynthetically reacting the reactant gas with saidculture, from the gas mixture, said removing the molecular oxygencomprising enabling a combustion reaction with the molecular oxygen andat least one other element to form a stable chemical compound thereofwith at least one other element, and without the stable chemicalcompound or other by-product of the chemical reaction being recirculatedinto said algal culture; and (d) recirculating the unconsumed reactantgas after oxygen removal into said algal culture in said substantiallysealed environment.
 18. The method of performing a continuousphotosynthetic reaction process as set forth in claim 17 wherein saidremoving said molecular oxygen comprises reacting said molecular oxygenwith hydrogen in a catalytic converter to form water as at least oneby-product.
 19. The method of performing a continuous photosyntheticreaction process as set forth in claim 18 further comprising the step ofcontrolling the introduction of hydrogen into said catalytic converter.20. The method of performing a continuous photosynthetic reactionprocess as set forth in claim 19 further comprising sensing the level ofoxygen concentration in said reactant gas.
 21. The method of performinga continuous photosynthetic reaction process as set forth in claim 15 orclaim 20 wherein said step of controlling the addition of hydrogencomprises actuating a valve means responsive to said sensing the levelof oxygen concentration when a preset level of oxygen concentration isexceeded.
 22. A closed photobioreactor comprising:(a) container meansfor containing a photosynthetic algal culture in a substantially sealedenvironment; (b) means for introducing light into said container means;(c) means for introducing a reactant gas into said container means forphotosynthetically reacting with culture in said container means in thepresence of light such that molecular oxygen is produced as aby-product, the molecular oxygen and unconsumed reactant gas forming agas mixture; (d) means for removing the molecular oxygen from the gasmixture without leaving by-products of the oxygen removal to berecirculated and without substantial leakage of the unconsumed reactantgas; (e) means for recirculating the unconsumed reactant gas afteroxygen removal through an algal culture in the substantially sealedenvironment; and wherein said means for removing molecular oxygencomprises means for enabling the molecular oxygen to react with hydrogenthrough an ion exchange membrane.
 23. The closed photobioreactor as setforth in claim 22, wherein said means for enabling comprises a firstchamber for containing the molecular oxygen, a second chamber forcontaining hydrogen and an ion exchange membrane separating said firstand second chambers for enabling the hydrogen when present in saidsecond chamber to react with said oxygen.